Development of laboratory procedures for handling a variety of sample types. Establishment of the NIH Bioinformatics Cooperative (http://affylims.cit.nih.gov/); development and dissemination of the MSCL toolbox; construction of a microarray database. Co-founded the NIH sponsored Symposium on the Functional Genomics of Critical Illness and Injury (last meeting: December 2009). This meeting led to the formation of the US Critical Illness and Injury Trials Group (USCIITG: http://www.usciitg.org/). Interactions between IFN-alpha and dexamethasone were investigated in primary bronchial epithelial cells (Physiologic Genomics 2005). Global transcriptional analysis of circulating leukocytes demonstrated oxidant and inflammatory vascular injury in sickle cell disease (Blood 2004). The human reticulocyte transcriptome was characterized extending our understanding of circulating cell-types (Physiol Genomics 2007). Expression profiling of platelets in sickle cell disease associated circulatory stress in this disorder with abnormalities of arginine metabolism (Circulation 2007). Single dose intravenous endotoxin administration to humans produced a rich profile of gene expression changes in blood (Physiol Genomics 2006). T lymphocyte-associated genes were repressed. Notably, these alterations in gene expression were rapidly extinguished within 24 h. Gene expression profiling was used to globally identify numerous genes regulated by NO. This work led to the discovery of new signal transduction pathways and regulatory mechanisms by which NO influences inflammation and cell proliferation (BMC Genomics 2005; Nuc Acid Res 2006; J Biol Chem 2006). A germline activating mutation in NRAS was found to cause autoimmune lymphoproliferative syndrome (PNAS 2007), suggesting that RAS-inactivating drugs might be therapeutically useful in autoimmune disorders. The innate and adaptive immune response to Pneumocystis was characterized in wild type and CD40-ligand knockout mice (J Leukoc Biol 2008). A focused mitochondria/metabolism/inflammation microarray was used to study HIV-associated myopathy/fatigue (Biol Res Nurs 2008; J Infect Dis 2012; Biol Res Nurs 2013). Genome-wide microarrays uncovered the importance of leptin and the thermogenic glycerolipid/fatty acid cycling pathway in protecting the heart from diet-induced steatosis (Physiological Genomics 2011). Microarrays were also used to globally examine the ability of carbon monoxide, an endogenous messenger produced by heme oxygenase, to suppress LPS-mediated gene induction in human monocytes (PLoS One 2009). Other studies examined endothelial progenitor cells (Arteriosclerosis Thrombosis and Vascular Biology 2009), animal strain effects on monocyte gene expression in a rat model of acute cardiac rejection (BMC Genomics 2009), and differences between CD4+ and CD8+ effector T cells in antitumor immunity (PNAS 2009). Targeted studies of specific inflammatory responses investigated gene regulation mediated by leukotriene D(4) activation of the type I cysteinyl leukotriene receptor (J Allergy Clin Immunol 2008) and the redundancy of leukotriene B(4) and D(4) signaling (Allergy 2011) in human monocytes. In air-liquid interface differentiated normal human bronchial epithelial cells challenged with IFN, O2 (70%) and NO (40 ppm), two gases used in the management of ARDS, produced highly similar signatures of oxidant stress (American Thoracic Society abstract 2011). Specific miRNAs in human airway epithelial cells were found to control gene and protein expression important for differentiation (Am J Respir Cell Mol Biol 2013). Dendritic cells (DCs) comprise distinct populations with specialized immune-regulatory functions. Retinoic acid (RA), a vitamin A derivative, was found to control the homeostasis of dendritic cell (DC) precursors. Total body radiation in patients and mice led to a selective loss of RA-dependent DC subsets and impaired class II restricted auto and antitumor immunity that could be rescued by supplemental RA (J Exp Med 2013). Global gene-expression changes across multiple organs implicated a host-wide IFN-response in staphylococcal enterotoxin B (SEB)-induced death (PLoS One 2014). After cord blood/haploidentical transplantation, engraftment of donor cells with germ-line integration of HHV6 mimics HHV6 reactivation (Blood 2014). Circulating endothelial cells (CECs) were identified by flow cytometry and their endothelial phenotype was validated using ultramicro analytical immunochemistry. Most CECs in healthy individuals are senescent without an identifiable nucleus and lack sufficient RNA for transcriptomal analysis (Thrombosis and Haemostasis 2014). Loss-of-function mutations in BMPR2 are the most common cause of heritable PAH. BMPR2-silenced human pulmonary artery endothelial cells (PAECs) linked oncogenic Ras/Raf/ERK signaling to phenotypic changes including proliferation, invasiveness and cytoskeletal abnormalities. Therapeutics targeting non-canonical Ras/Raf/ERK signaling may be useful in preventing or reversing vascular remodeling in patients with PAH (Am J Physiol Lung Cell Mol Physiol. 2016). In human endothelial cells, MR agonists were found to repress NF-kB mediated gene transcription. In contrast, MR either repressed or further activated AP1 signaling depending on the DNA sequence of specific AP1 binding sites, MR ligand structure, and AP1 family member expression. Global expression profiling identified target genes that were trans-activated by this MR/AP1-dependent pro-inflammatory mechanism (J Biol Chem 2016). The comparative biology of the many genomic mutations associated with the development of PAH has the potential to identify both individualized and possibly universal therapeutic targets aimed at halting or even reversing the progression of pathologic vascular remodeling. Genome-wide expression profiling studies of BMPR2-, CAV1-, SMAD9, COUPTFII and PHD2-silenced primary human pulmonary artery endothelial cells are providing a comprehensive picture of the shared and divergent networks associated with PAH pathobiology. In addition, techniques are being developed to produce induced pluripotent stem cells (iPS) from PAH patient-derived fibroblasts and peripheral blood. Dermal fibroblasts from PAH patients with CAV1 mutations displayed an abnormal cellular phenotype that mirrored the phenotype we observed in CAV1-silenced PAECs. Thus, non-lung cells, such as dermal fibroblasts, harboring a PAH-associated mutation might be useful for investigating associated signaling abnormalities as well as potential therapeutic targets in this disease (ATS abstract 2017). CRISPR/CAS9 is also being explored as a means to create or repair specific PAH-associated genetic defects in selected cell types. Emerging areas of interest for better understanding PAH pathogenesis include oncogenic signaling, inflammation, endothelial to mesenchymal transition, and pseudohypoxia. Transcriptomic and morphologic analyses of heterotopic hearts in a rat cardiac transplant model demonstrated significant mitochondrial injury during acute cardiac rejection with downregulation of mitochondrial energetics (ISHLT abstract 2017). To better understand altered inflammatory responses in PAH, functional loss of BMPR2 is being investigated in human monocytes and macrophages, as well as the impact of pseudohypoxia on lymphocyte immunity.